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Sir, We read with interest the recent paper by Odds et al., as we have performed a similar study in which we measured the concentrations of itraconazole (IT) and its metabolite, hydroxy-itraconazole (HI), in the sera of 40 patients by a bioassay and compared the results with those determined by HPLC; in common with these investigators, we used IT and HI as standards. The findings of the two studies are similar, i.e. a bioassay in which IT is used as the standard overestimates the serum concentrations of the drug, whereas one in which HI is used underestimates the concentrations. In our study, the concentrations obtained with HI as the standard were approximately 50% of the combined IT and HI concentrations determined by HPLC and it was therefore necessary to apply a correction factor of two to the results determined by the bioassay. In the study reported by Odds et al., five experienced laboratories employed a bioassay technique with standards containing either HI or IT to measure the concentrations of itraconazole in 10 spiked samples. There was considerable interlaboratory variation, with mean values differing markedly from the target values (Table). With linear regression analysis, we examined the relationship between the bioassay and HPLC (target) results for each laboratory. For bioassays in which HI standards were used, the correction factors (slope) ranged from 1.04 to 5.1 (mean 2.05), whereas when IT standards were used, the correction factors ranged from 0.26 to 2.12 (mean 0.73). These data suggest that results obtained with a bioassay will differ from those obtained with HPLC, irrespective of whether HI or IT is used as the standard, and that interlaboratory variation is inevitable, even when the assays are undertaken in laboratories experienced in doing so, although the results are likely to be less reliable when assays are performed by inexperienced laboratory staff. The need to apply a correction factor to results obtained with a bioassay will probably be laboratory dependent and will be influenced markedly by the assay conditions. To enable laboratories to calculate their own correction factors, standards containing known concentrations of HI or IT should be assayed at regular intervals. An external quality assessment scheme that allows laboratories to monitor both their performances and correction factors would also be advantageous. The present study and our earlier observations suggest that further research into assay methods is required urgently, to identify the factors that have the greatest effects on the results obtained with bioassays and to define

Discrepancies associated with the measurement of itraconazole serum concentrations by bioassays